Limited resources of fossil fuels, and increasing amounts of CO2 released from them and causing the greenhouse phenomenon have raised a need for using biomass as a renewable and clean source of energy. Biomass resources can be broadly categorized as agricultural or forestry-based, including secondary sources derived from agro and wood industries, waste sources and municipal solid wastes. One promising, alternative technology is the production of biofuels i.e. (bio)ethanol from lignocellulosic materials. In the transportation sector biofuels are for the time being the only option, which could reduce the CO2 emissions by an order of magnitude. The ethanol can be used in existing vehicles and distribution systems and thus it does not require expensive infrastructure investments. Sugars derived from lignocellulosic renewable raw materials can also be used as raw materials for a variety of chemical products that can replace oil-based chemicals.
Lignocellulosic raw material comprises an abundant source of carbohydrates for a variety of biofuels, including bioethanol. Most of the carbohydrates in plants are in the form of lignocellulose, which essentially consists of cellulose, hemicellulose, and lignin. Lignocellulose can be converted into bioethanol and other chemical products via fermentation following hydrolysis to fermentable sugars. In a conventional lignocellulose-to-ethanol process the lignocellulosic material is first pretreated either chemically or physically to make the cellulose fraction more accessible to hydrolysis. The cellulose fraction is then hydrolysed to obtain sugars that can be fermented by yeast into ethanol and distilled to obtain pure ethanol. Lignin is obtained as a main co-product that may be used as a solid fuel.
One barrier of production of biofuels from cellulosic and lignocellulosic biomass is the robustness of the cell walls and the presence of sugar monomers in the form of inaccessible polymers that require a great amount of processing to make sugar monomers available to the micro-organisms that are typically used to produce alcohol by fermentation. Enzymatic hydrolysis is considered the most promising technology for converting cellulosic biomass into fermentable sugars. However, enzymatic hydrolysis is used only to a limited amount at industrial scale, and especially when using strongly lignified material such as wood or agricultural waste the technology is not satisfactory. The cost of the enzymatic step is one of the major economic factors of the process. Efforts have been made to improve the efficiency of the enzymatic hydrolysis of the cellulosic material (Badger 2002).
WO2001060752 describes a continuous process for converting solid lignocellulosic biomass into combustible fuel products. After pretreatment by wet oxidation or steam explosion the biomass is partially separated into cellulose, hemicellulose and lignin, and is then subjected to partial hydrolysis using one or more carbohydrase enzymes (EC 3.2).
WO2002024882 concerns a method of converting cellulose to glucose by treating a pretreated lignocellulosic substrate with an enzyme mixture comprising cellulase and a modified cellobiohydrolase I (CBHI) obtained by inactivating its cellulose binding domain (CBD).
US 20040005674 A1 describes novel enzyme mixtures that can be used directly on lignocellulose substrate, whereby toxic waste products formed during pretreatment processes may be avoided, and energy may be saved. The synergistic enzyme mixture contains a cellulase and an auxiliary enzyme such as xylanase, ligninase, amylase, protease, lipidase or glucuronidase, or any combination thereof. Cellulase is considered to include endoglucanase, beta-glucosidase and cellobiohydrolase. US 20050164355 describes a method for degrading lignocellulosic material with one or more cellulolytic enzymes selected from endoglucanase, beta-glucosidase and cellobiohydrolase and in the presence of at least one surfactant. Additional enzymes such as hemicellulases, esterase, peroxidase, protease, laccase or mixture thereof may also be used. The presence of surfactant increases the degradation of lignocellulosic material compared to the absence of surfactant.
WO2011080317 describes a method of treating cellulosic material with fungal CBHII/Cel6A cellobiohydrolase enzyme. The enzyme is useful in various industrial applications, particularly in production of biofuels, where production of fermentable sugars from lignocellulosic material at moderate to elevated temperature is advantageous.
Cellulases from a number of bacterial and fungal sources have been purified and characterized. The best investigated and most widely applied cellulolytic enzymes of fungal origin have been derived from Trichoderma reesei (the anamorph of Hypocrea jecorina). Cellulases from less known fungi have also been disclosed. Hong et al. (2003a and 2003b) characterize EG and CBHI of Thermoascus aurantiacus produced in yeast. Tuohy et al. (2002) describe three forms of cellobiohydrolases from Talaromyces emersonii, a moderately thermophilic fungus. The sequence and detailed biochemical characterization of these T. emersonii cellobiohydrolases have shown comparable properties with the cellobiohydrolases of T. reesei and P. chrysosporium. The cellulase enzymes of another thermophilic fungus, Melanocarpus albomyces, include at least two endoglucanases (Cel45A and Cel7A) and one cellobiohydrolase (Cel7B). These enzymes have been cloned and characterized for their pH and temperature behavior (Miettinen-Oinonen et al., 2004). WO2007071818 describes enzymatic conversion of lignocellulosic material by enzymes including cellobiohydrolase, endoglucanase, beta-glucosidase and optionally xylanase derived from Thermoascus auranticus, Acremonium thermophilium or Chaetomium thermophilium. U.S. Pat. No. 7,892,812 describes cellulose compositions comprising endoglucanase and their use in industrial applications, for example in saccharification of lignocellulose biomass. The cellulases are from fungi Chrysosporium lucknowense, which has been identified as Myceliophthora thermophila (Visser et al., 2011).
Endoglucanases of the Cel7 family (EGs fam 7) are disclosed e.g. in U.S. Pat. No. 5,912,157, which pertains Myceliphthora endoglucanase and its homologues and applications thereof in detergent, textile, and pulp. U.S. Pat. No. 6,071,735 describes cellulases exhibiting high endoglucanase activity in alkaline conditions. Uses as detergent, in pulp and paper, and textile applications are discussed. U.S. Pat. No. 5,763,254 discloses enzymes from strains of Humicola, Fusarium and Myceliopthora degrading cellulose/hemicellulose and having a carbohydrate binding module homologous to the region A of T. reesei. 
WO2004078919 discloses purified glycosyl hydrolase family 7 (Cel7A) enzymes from Penicillium funiculosum, which demonstrate a high level of specific performance when formulated with an endoglucanase and tested on pretreated corn stover.
Haakana et al., (2004) describes the cloning and sequencing of three genes encoding cellulases Cel45A, Cel7A and Cel7B from Melanocarpus albomyces. These cellulases work well in biostoning, with lower backstaining compared to T. reesei. WO9714804 discloses Cel7A family enzymes from Melanocarpus albomyces and its applications in textile and detergent industry. Voutilainen et al., (2008) describes novel GH7 family cellobiohydrolases from the thermophilic fungi Acremonium thermophilum, Thermoascus auranticus and Chaetomium thermophilum active on insoluble polymeric substrates and participating in the rate limiting step in the hydrolysis of cellulose.
U.S. Pat. No. 5,393,670 describes the DNA, vectors and transformed host encoding Trichoderma reesei endoglucanase I.
There is a continuous need for new methods of degrading cellulosic substrates, in particular lignocellulosic substrates, and for new enzymes and enzyme mixtures, which enhance the efficiency of the degradation. There is also a need for enzymes and processes, which are versatile and which work not only at moderate temperatures but also at high temperatures, thus increasing the reaction rates and enabling the use of high biomass consistency leading to high sugar and ethanol concentrations. This approach may lead to significant savings in energy and investment costs. The high temperature also decreases the risk of contamination during hydrolysis. The present invention aims to meet at least part of these needs.